Obstructive sleep apnea in a mouse model is associated with tissue-specific transcriptomic changes in circadian rhythmicity and mean 24-hour gene expression.

Intermittent hypoxia (IH) is a major clinical feature of obstructive sleep apnea (OSA). The mechanisms that become dysregulated after periods of exposure to IH are unclear, particularly in the early stages of disease. The circadian clock governs a wide array of biological functions and is intimately associated with stabilization of hypoxia-inducible factors (HIFs) under hypoxic conditions. In patients, IH occurs during the sleep phase of the 24-hour sleep-wake cycle, potentially affecting their circadian rhythms. Alterations in the circadian clock have the potential to accelerate pathological processes, including other comorbid conditions that can be associated with chronic, untreated OSA. We hypothesized that changes in the circadian clock would manifest differently in those organs and systems known to be impacted by OSA. Using an IH model to represent OSA, we evaluated circadian rhythmicity and mean 24-hour expression of the transcriptome in 6 different mouse tissues, including the liver, lung, kidney, muscle, heart, and cerebellum, after a 7-day exposure to IH. We found that transcriptomic changes within cardiopulmonary tissues were more affected by IH than other tissues. Also, IH exposure resulted in an overall increase in core body temperature. Our findings demonstrate a relationship between early exposure to IH and changes in specific physiological outcomes. This study provides insight into the early pathophysiological mechanisms associated with IH.

Time-restricted feeding protects the blood pressure circadian rhythm in diabetic mice.

The quantity and quality of food intake have been considered crucial for peoples' wellness. Only recently has it become appreciated that the timing of food intake is also critical. Nondipping blood pressure (BP) is prevalent in diabetic patients and is associated with increased cardiovascular events. However, the causes and mechanisms of nondipping BP in diabetes are not fully understood. Here, we report that food intake and BP were arrhythmic in diabetic db/db mice fed a normal chow diet ad libitum. Imposing a food intake diurnal rhythm by time-restricted feeding (TRF; food was only available for 8 h during the active phase) prevented db/db mice from developing nondipping BP and effectively restored the already disrupted BP circadian rhythm in db/db mice. Interestingly, increasing the time of food availability from 8 h to 12 h during the active dark phase in db/db mice prompted isocaloric feeding and still provided robust protection of the BP circadian rhythm in db/db mice. In contrast, neither 8-h nor 12-h TRF affected BP dipping in wild-type mice. Mechanistically, we demonstrate that TRF protects the BP circadian rhythm in db/db mice via suppressing the sympathetic activity during the light phase when they are inactive and fasting. Collectively, these data reveal a potentially pivotal role of the timing of food intake in the prevention and treatment of nondipping BP in diabetes.

Authentication of In Situ Measurements for Thoracic Aortic Aneurysms in Mice.

[Figure: see text].

Myocardial-restricted ablation of the GTPase RAD results in a pro-adaptive heart response in mice.

Existing therapies to improve heart function target ß-adrenergic receptor (ß-AR) signaling and Ca2+ handling and often lead to adverse outcomes. This underscores an unmet need for positive inotropes that improve heart function without any adverse effects. The GTPase Ras associated with diabetes (RAD) regulates L-type Ca2+ channel (LTCC) current (ICa,L). Global RAD-knockout mice (gRAD-/-) have elevated Ca2+ handling and increased cardiac hypertrophy, but RAD is expressed also in noncardiac tissues, suggesting the possibility that pathological remodeling is due also to noncardiac effects. Here, we engineered a myocardial-restricted inducible RAD-knockout mouse (RADΔ/Δ). Using an array of methods and techniques, including single-cell electrophysiological and calcium transient recordings, echocardiography, and radiotelemetry monitoring, we found that RAD deficiency results in a sustained increase of inotropy without structural or functional remodeling of the heart. ICa,L was significantly increased, with RAD loss conferring a ß-AR-modulated phenotype on basal ICa,L Cardiomyocytes from RADΔ/Δ hearts exhibited enhanced cytosolic Ca2+ handling, increased contractile function, elevated sarcoplasmic/endoplasmic reticulum calcium ATPase 2 (SERCA2a) expression, and faster lusitropy. These results argue that myocardial RAD ablation promotes a beneficial elevation in Ca2+ dynamics, which would obviate a need for increased ß-AR signaling to improve cardiac function.

Deletion of BMAL1 in Smooth Muscle Cells Protects Mice From Abdominal Aortic Aneurysms.

OBJECTIVE: Abdominal aortic aneurysm (AAA) has high mortality rate when ruptured, but currently, there is no proven pharmacological therapy for AAA because of our poor understanding of its pathogenesis. The current study explored a novel role of smooth muscle cell (SMC) BMAL1 (brain and muscle Arnt-like protein-1)-a transcription factor known to regulate circadian rhythm-in AAA development. APPROACH AND RESULTS: SMC-selective deletion of BMAL1 potently protected mice from AAA induced by (1) MR (mineralocorticoid receptor) agonist deoxycorticosterone acetate or aldosterone plus high salt intake and (2) angiotensin II infusion in hypercholesterolemia mice. Aortic BMAL1 was upregulated by deoxycorticosterone acetate-salt, and deletion of BMAL1 in SMCs selectively upregulated TIMP4 (tissue inhibitor of metalloproteinase 4) and suppressed deoxycorticosterone acetate-salt-induced MMP (matrix metalloproteinase) activation and elastin breakages. Moreover, BMAL1 bound to the Timp4 promoter and suppressed Timp4 transcription. CONCLUSIONS: These results reveal an important, but previously unexplored, role of SMC BMAL1 in AAA. Moreover, these results identify TIMP4 as a novel target of BMAL1, which may mediate the AAA protective effect of SMC BMAL1 deletion.

Mineralocorticoid receptor agonists induce mouse aortic aneurysm formation and rupture in the presence of high salt.

OBJECTIVE: Elevated plasma aldosterone concentrations in patients have been linked to a spectrum of cardiovascular diseases. Mineralocorticoid receptor antagonists provide additional benefits in patients with heart failure. However, whether aldosterone and the mineralocorticoid receptor are involved in aortic aneurysm is unknown. APPROACH AND RESULTS: We report that administration of deoxycorticosterone acetate (DOCA) and salt or aldosterone and salt, but not DOCA or salt alone, to C57BL/6 male mice induced abdominal and thoracic aortic aneurysm formation and rupture in an age-dependent manner. DOCA and salt- or aldosterone and salt-induced aortic aneurysm mimicked human aortic aneurysm with respect to elastin degradation, inflammatory cell infiltration, smooth muscle cell degeneration and apoptosis, and oxidative stress. Aortic aneurysm formation did not correlate with the increase in blood pressure induced by DOCA and salt. Systemic administration of the angiotensin-converting enzyme inhibitor, enalapril, or angiotensin type 1 receptor antagonist, losartan, did not affect DOCA and salt-induced aortic aneurysm. In contrast, the mineralocorticoid receptor antagonists, spironolactone or eplerenone, significantly attenuated DOCA and salt- or aldosterone and salt-induced aortic aneurysm. CONCLUSIONS: The current study describes a novel aortic aneurysm animal model induced by mineralocorticoid receptor agonist and high salt, and reveals a previously unrecognized but potentially significant role of aldosterone in the pathogenesis of aortic aneurysm. These findings imply that mineralocorticoid receptor antagonists may be effective in the treatment of some aortic aneurysms.

Exenatide administration time determines the effects on blood pressure dipping in db/db mice via modulation of food intake and sympathetic activity.

Type 2 diabetics have an increased prevalence of hypertension and nondipping blood pressure (BP), which worsen cardiovascular outcomes. Exenatide, a short acting glucagon-like peptide-1 receptor agonist (GLP-1RA) used to treat type 2 diabetes, also demonstrates blood pressure (BP)-lowering effects. However, the mechanisms behind this and the impact of administration timing on BP dipping remain unclear. We investigated the effects of exenatide intraperitoneal injected at light onset (ZT0) or dark onset (ZT12) in diabetic (db/db) mice and nondiabetic controls. Using radio-telemetry and BioDAQ cages, we continuously monitored BP and food intake. Db/db mice exhibited non-dipping BP and increased food intake. ZT0 exenatide administration restored BP dipping by specifically lowering light-phase BP, while ZT12 exenatide reversed dipping by lowering dark-phase BP. These effects correlated with altered food intake patterns, and importantly, were abolished when food access was removed. Additionally, urinary norepinephrine excretion, measured by HPLC, was significantly reduced 6 hours post-exenatide at both ZT0 and ZT12, suggesting sympathetic nervous system involvement. Notably, combining exenatide with either ganglionic blocker mecamylamine or α-blocker prazosin did not enhance BP reduction beyond the individual effects of each blocker. These findings reveal that exenatide, when administered at light onset, restores BP dipping in db/db mice by suppressing light-phase food intake and sympathetic activity. Importantly, the efficacy of exenatide is dependent on food availability and its timing relative to circadian rhythms, highlighting the potential for chronotherapy in optimizing GLP-1RA- based treatments for type 2 diabetes and hypertension. Article Highlights: Maintaining a normal blood pressure (BP) circadian rhythm is vital for cardiovascular health, but diabetes often disrupts this rhythm. The effect of exenatide, a GLP-1 receptor agonist (GLP-1RA), on BP rhythm in diabetes is uncertain.This study investigates the impact of exenatide administration timing on BP patterns in diabetic db/db mice.Findings indicate that exenatide given at the onset of rest restores normal BP dipping, while at the start of the active phase worsens BP rhythm by modulating food intake and sympathetic activity.Timing GLP-1 RA administration may optimize BP control and provide cardiovascular benefits for type 2 diabetes patients.

Androgen aggravates aortic aneurysms via suppression of PD-1 in mice.

Androgen has long been recognized for its pivotal role in the sexual dimorphism of cardiovascular diseases, including aortic aneurysms (AAs), a devastating vascular disease with a higher prevalence and fatality rate in men than in women. However, the mechanism by which androgen mediates AAs is largely unknown. Here, we found that male, not female, mice developed AAs when exposed to aldosterone and high salt (Aldo-salt). We revealed that androgen and androgen receptors (ARs) were crucial for this sexually dimorphic response to Aldo-salt. We identified programmed cell death protein 1 (PD-1), an immune checkpoint, as a key link between androgen and AAs. Furthermore, we demonstrated that administration of anti-PD-1 Ab and adoptive PD-1-deficient T cell transfer reinstated Aldo-salt-induced AAs in orchiectomized mice and that genetic deletion of PD-1 exacerbated AAs induced by a high-fat diet and angiotensin II (Ang II) in nonorchiectomized mice. Mechanistically, we discovered that the AR bound to the PD-1 promoter to suppress the expression of PD-1 in the spleen. Thus, our study unveils a mechanism by which androgen aggravates AAs by suppressing PD-1 expression in T cells. Moreover, our study suggests that some patients with cancer might benefit from screenings for AAs during immune checkpoint therapy.

Smooth muscle-specific expression of calcium-independent phospholipase A2ß (iPLA2ß) participates in the initiation and early progression of vascular inflammation and neointima formation.

Whether group VIA phospholipase A(2) (iPLA(2)ß) is involved in vascular inflammation and neointima formation is largely unknown. Here, we report that iPLA(2)ß expression increases in the vascular tunica media upon carotid artery ligation and that neointima formation is suppressed by genetic deletion of iPLA(2)ß or by inhibiting its activity or expression via perivascular delivery of bromoenol lactone or of antisense oligonucleotides, respectively. To investigate whether smooth muscle-specific iPLA(2)ß is involved in neointima formation, we generated transgenic mice in which iPLA(2)ß is expressed specifically in smooth muscle cells and demonstrate that smooth muscle-specific expression of iPLA(2)ß exacerbates ligation-induced neointima formation and enhanced both production of proinflammatory cytokines and vascular infiltration by macrophages. With cultured vascular smooth muscle cell, angiotensin II, arachidonic acid, and TNF-α markedly induce increased expression of IL-6 and TNF-α mRNAs, all of which were suppressed by inhibiting iPLA(2)ß activity or expression with bromoenol lactone, antisense oligonucleotides, and genetic deletion, respectively. Similar suppression also results from genetic deletion of 12/15-lipoxygenase or inhibiting its activity with nordihydroguaiaretic acid or luteolin. Expression of iPLA(2)ß protein in cultured vascular smooth muscle cells was found to depend on the phenotypic state and to rise upon incubation with TNF-α. Our studies thus illustrate that smooth muscle cell-specific iPLA(2)ß participates in the initiation and early progression of vascular inflammation and neointima formation and suggest that iPLA(2)ß may represent a novel therapeutic target for preventing cardiovascular diseases.

Smooth muscle-selective CPI-17 expression increases vascular smooth muscle contraction and blood pressure.

Recent data revealed that protein kinase C-potentiated myosin phosphatase inhibitor of 17 kDa (CPI-17), a myosin phosphatase inhibitory protein preferentially expressed in smooth muscle, is upregulated/activated in several diseases but whether this CPI-17 increase plays a causal role in pathologically enhanced vascular smooth muscle contractility and blood pressure remains unclear. To address this possibility, we generated a smooth muscle-specific CPI-17 transgenic mouse model (CPI-17-Tg) and demonstrated that the CPI-17 transgene was selectively expressed in smooth muscle-enriched tissues, including mesenteric arteries. The isometric contractions in the isolated second-order branch of mesenteric artery helical strips from CPI-17-Tg mice were significantly enhanced compared with controls in response to phenylephrine, U-46619, serotonin, ANG II, high potassium, and calcium. The perfusion pressure increases in isolated perfused mesenteric vascular beds in response to norepinephrine were also enhanced in CPI-17-Tg mice. The hypercontractility was associated with increased phosphorylation of CPI-17 and 20-kDa myosin light chain under basal and stimulated conditions. Surprisingly, the protein levels of rho kinase 2 and protein kinase Cα/δ were significantly increased in CPI-17-Tg mouse mesenteric arteries. Radiotelemetry measurements demonstrated that blood pressure was significantly increased in CPI-17-Tg mice. However, no vascular remodeling was detected by morphometric analysis. Taken together, our results demonstrate that increased CPI-17 expression in smooth muscle promotes vascular smooth muscle contractility and increases blood pressure, implicating a pathological significant role of CPI-17 upregulation.

Androgen aggravates aortic aneurysms via suppressing PD-1 in mice.

Androgen has long been recognized for its pivotal role in the sexual dimorphism of cardiovascular diseases, including aortic aneurysms, a devastating vascular disease with a higher prevalence and mortality rate in men than women. However, the molecular mechanism by which androgen mediates aortic aneurysms is largely unknown. Here, we report that male but not female mice develop aortic aneurysms in response to aldosterone and high salt (Aldo-salt). We demonstrate that both androgen and androgen receptors (AR) are crucial for the sexually dimorphic response to Aldo-salt. We identify T cells expressing programmed cell death protein 1 (PD-1), an immune checkpoint molecule important in immunity and cancer immunotherapy, as a key link between androgen and aortic aneurysms. We show that intraperitoneal injection of anti-PD-1 antibody reinstates Aldo-salt-induced aortic aneurysms in orchiectomized mice. Mechanistically, we demonstrate that AR binds to the PD-1 promoter to suppress its expression in the spleen. Hence, our study reveals an important but unexplored mechanism by which androgen contributes to aortic aneurysms by suppressing PD-1 expression in T cells. Our study also suggests that cancer patients predisposed to the risk factors of aortic aneurysms may be advised to screen for aortic aneurysms during immune checkpoint therapy.

Prenatal Morphine Exposure Increases Cardiovascular Disease Risk and Programs Neurogenic Hypertension in the Adult Offspring.

BACKGROUND: The opioid overdose and opioid use disorder epidemics are concomitant with increased metabolic and CVD risk. Although opioid use disorder causes adverse pregnancy outcomes, the offspring's cardiovascular health is understudied. We hypothesized that offspring exposed to in utero morphine exposure (IUME) would show increased CVD risk factors and endogenous opioid system dysregulation. METHODS: Sprague Dawley dams were treated with saline (vehicle, n=10) or escalating doses of morphine (5-20 mg/kg per day, SC, n=10) during gestation. Cardiovascular and metabolic parameters were assessed in adult offspring. RESULTS: Litter size and pups' birth weight were not different in response to IUME. Female and male IUME offspring showed reduced body length at birth (P<0.05) and body weight from weeks 1 to 3 of life (P<0.05), followed by a catch-up growth effect. By week 16, female and male IUME rats showed reduced tibia length (P<0.05) and fat mass. IUME increases the mean arterial pressure and the depressor response to mecamylamine (5 mg/kg per day, IP) induced by IUME were abolished by a chronic treatment with an alpha-adrenergic receptor blocker (prazosin; 1 mg/kg per day, IP). Although circulating levels of angiotensin peptides were similar between groups, IUME exacerbated maximal ex vivo Ang (angiotensin) II-induced vasoconstriction (P<0.05) and induced endothelial dysfunction in a sex-specific manner (P<0.05). Proenkephalin, an endogenous opioid peptide that lowers blood pressure and sympathetic-mediated vasoconstriction, showed reduced mRNA expression in the heart, aorta, and kidneys from morphine versus vehicle group (P<0.05). CONCLUSIONS: Among the effects of IUME, neurogenic hypertension, vascular dysfunction, and metabolic dysfunction could be associated with the dysregulation of the endogenous opioid system.

Toward Precision Medicine: Circadian Rhythm of Blood Pressure and Chronotherapy for Hypertension - 2021 NHLBI Workshop Report.

Healthy individuals exhibit blood pressure variation over a 24-hour period with higher blood pressure during wakefulness and lower blood pressure during sleep. Loss or disruption of the blood pressure circadian rhythm has been linked to adverse health outcomes, for example, cardiovascular disease, dementia, and chronic kidney disease. However, the current diagnostic and therapeutic approaches lack sufficient attention to the circadian rhythmicity of blood pressure. Sleep patterns, hormone release, eating habits, digestion, body temperature, renal and cardiovascular function, and other important host functions as well as gut microbiota exhibit circadian rhythms, and influence circadian rhythms of blood pressure. Potential benefits of nonpharmacologic interventions such as meal timing, and pharmacologic chronotherapeutic interventions, such as the bedtime administration of antihypertensive medications, have recently been suggested in some studies. However, the mechanisms underlying circadian rhythm-mediated blood pressure regulation and the efficacy of chronotherapy in hypertension remain unclear. This review summarizes the results of the National Heart, Lung, and Blood Institute workshop convened on October 27 to 29, 2021 to assess knowledge gaps and research opportunities in the study of circadian rhythm of blood pressure and chronotherapy for hypertension.

Altered clock gene expression and vascular smooth muscle diurnal contractile variations in type 2 diabetic db/db mice.

This study was designed to determine whether the 24-h rhythms of clock gene expression and vascular smooth muscle (VSM) contractile responses are altered in type 2 diabetic db/db mice. Control and db/db mice were euthanized at 6-h intervals throughout the day. The aorta, mesenteric arteries, heart, kidney, and brain were isolated. Clock and target gene mRNA levels were determined by either real-time PCR or in situ hybridization. Isometric contractions were measured in isolated aortic helical strips, and pressor responses to an intravenous injection of vasoconstrictors were determined in vivo using radiotelemetry. We found that the 24-h mRNA rhythms of the following genes were suppressed in db/db mice compared with control mice: the clock genes period homolog 1/2 (Per1/2) and cryptochrome 1/2 (Cry1/2) and their target genes D site albumin promoter-binding protein (Dbp) and peroxisome proliferator-activated receptor-γ (Pparg) in the aorta and mesenteric arteries; Dbp in the heart; Per1, nuclear receptor subfamily 1, group D, member 1 (Rev-erba), and Dbp in the kidney; and Per1 in the suprachiasmatic nucleus. The 24-h contractile variations in response to phenylephrine (α(1)-agonist), ANG II, and high K(+) were significantly altered in the aortas from db/db mice compared with control mice. The diurnal variations of the in vivo pressor responses to phenylephrine and ANG II were lost in db/db mice. Moreover, the 24-h mRNA rhythms of the contraction-related proteins Rho kinase 1/2, PKC-potentiated phosphatase inhibitory protein of 17 kDa, calponin-3, tropomyosin-1/2, and smooth muscle protein 22-α were suppressed in db/db mice compared with control mice. Together, our data demonstrated that the 24-h rhythms of clock gene mRNA, mRNA levels of several contraction-related proteins, and VSM contraction were disrupted in db/db mice, which may contribute to the disruption of their blood pressure circadian rhythm.

Role of sympathetic pathway in light-phase time-restricted feeding-induced blood pressure circadian rhythm alteration.

Disruption of blood pressure (BP) circadian rhythm, independent of hypertension, is emerging as an index for future target organ damage and is associated with a higher risk of cardiovascular events. Previous studies showed that changing food availability time alters BP rhythm in several mammalian species. However, the underlying mechanisms remain largely unknown. To address this, the current study specifically investigates (1) the relationship between rhythms of food intake and BP in wild-type mice; (2) effects of light-phase time-restricted feeding (TRF, food only available during light-phase) on BP circadian rhythm in wild-type and diabetic db/db mice; (3) the roles of the autonomic system and clock gene in light-phase TRF induced changes in BP circadian rhythm. Food intake and BP of C57BL/6J and db/db mice were simultaneously and continuously recorded using BioDAQ and telemetry systems under ad libitum or light-phase TRF. Per2 protein daily oscillation was recorded in vivo by IVIS spectrum in mPer2 Luc mice. Autonomic nerve activity was evaluated by heart rate variability, baroreflex, urinary norepinephrine (NE) and epinephrine (Epi) excretion, and mRNA expressions of catecholamines biosynthetic and catabolic enzymes, and alpha-adrenergic receptors in mesenteric resistance arteries. We found that in wild-type mice, the BP level was correlated with the food intake temporally across the 24 h. Reversing the feeding time by imposing light-phase TRF resulted in reverse or inverted BP dipping. Interestingly, the net changes in food intake were correlated with the net alteration in BP temporally under light-phase TRF. In db/db mice, light-phase TRF worsened the existing non-dipping BP. The food intake and BP circadian rhythm changes were associated with alterations in Per2 protein daily oscillation and the time-of-day variations in heart rate variability, baroreflex, and urinary excretion of NE and Epi, and increased mRNA expression of Slc6a2 (encoding NE transporter) and Adra1d (encoding alpha-adrenergic receptor 1d) in the mesenteric resistance arteries, indicating the sympathetic nervous system (SNS) was modulated after light-phase TRF. Collectively, our results demonstrated that light-phase TRF results in reverse dipping of BP in wild-type and diabetic db/db mice and revealed the potential role of the sympathetic pathway in light-phase TRF-induced BP circadian rhythm alteration.

Role of calcium-independent phospholipase A2beta in high glucose-induced activation of RhoA, Rho kinase, and CPI-17 in cultured vascular smooth muscle cells and vascular smooth muscle hypercontractility in diabetic animals.

Previous studies suggest that high glucose-induced RhoA/Rho kinase/CPI-17 activation is involved in diabetes-associated vascular smooth muscle hypercontractility. However, the upstream signaling that links high glucose and RhoA/Rho kinase/CPI-17 activation is unknown. Here we report that calcium-independent phospholipase A(2)beta (iPLA(2)beta) is required for high glucose-induced RhoA/Rho kinase/CPI-17 activation and thereby contributes to diabetes-associated vascular smooth muscle hypercontractility. We demonstrate that high glucose increases iPLA(2)beta mRNA, protein, and iPLA(2) activity in a time-dependent manner. Protein kinase C is involved in high glucose-induced iPLA(2)beta protein up-regulation. Inhibiting iPLA(2)beta activity with bromoenol lactone or preventing its expression by genetic deletion abolishes high glucose-induced RhoA/Rho kinase/CPI-17 activation, and restoring expression of iPLA(2)beta in iPLA(2)beta-deficient cells also restores high glucose-induced CPI-17 phosphorylation. Pharmacological and genetic inhibition of 12/15-lipoxygenases has effects on high glucose-induced CPI-17 phosphorylation similar to iPLA(2)beta inhibition. Moreover, increases in iPLA(2) activity and iPLA(2)beta protein expression are also observed in both type 1 and type 2 diabetic vasculature. Pharmacological and genetic inhibition of iPLA(2)beta, but not iPLA(2)gamma, diminishes diabetes-associated vascular smooth muscle hypercontractility. In summary, our results reveal a novel mechanism by which high glucose-induced, protein kinase C-mediated iPLA(2)beta up-regulation activates the RhoA/Rho kinase/CPI-17 via 12/15-lipoxygenases and thereby contributes to diabetes-associated vascular smooth muscle hypercontractility.

Circadian variations of vasoconstriction and blood pressure in physiology and diabetes.

The intrinsic vascular smooth muscle contraction and vasoconstriction show time-of-day variations, contributing to the blood pressure circadian rhythm, which is essential for cardiovascular health. This brief review provides an overview of our current understanding of the mechanisms underlying the time-of-day variations of vascular smooth muscle contraction. We discuss the potential contribution of the time-of-day variations of vasoconstriction to the physiological blood pressure circadian rhythm. Finally, we survey the data obtained in the type 2 diabetic db/db mouse model that demonstrate the alterations of the time-of-day variations of vasoconstriction and the nondipping blood pressure in diabetes.

Production and binding of endothelin-2 (EDN2) in the rat ovary: endothelin receptor subtype A (EDNRA)-mediated contraction.

Endothelin-2 (EDN2)-mediated contraction has been proposed as a final mechanical signal facilitating ovulation. The objectives herein were to determine (1) whether ovarian endothelins were increased before ovulation; (2) whether a specific endothelin-converting enzyme (ECE) was mediating their production; (3) which receptor was facilitating ovarian contraction; and (4) whether receptor-specific antagonism affected ovulation. Follicular development was induced in immature rats with 10 IU pregnant mare serum gonadotrophin (PMSG) and the ovulatory cascade was initiated 48 h later with 10 IU human chorionic gonadotrophin (hCG). In Experiment 1, an immunoassay revealed that the ovarian concentration of endothelin peptide was increased 7-fold 12 h after hCG when compared with 48 h after PMSG (P < 0.05). In Experiment 2, real-time PCR indicated that mRNA for Ece1, but not Ece2, was increased in granulosa cells collected 12 h after hCG when compared with those collected before the ovulatory stimulus (P < 0.05). In Experiment 3, isometric tension analysis revealed that the contractile effect of EDN2 was mediated by endothelin receptor A (EDNRA), not B (EDNRB). In Experiment 4, no effect was observed on the rate of ovulation when rats were treated with an antagonist specific to EDNRA (BQ123) or EDNRB (BQ788), or when mice were treated with BQ123, BQ788 or BQ123 + BQ788. In conclusion, endothelin peptide is produced before ovulation and the contractile action of EDN2 within the ovary is facilitated via EDNRA. In addition, findings of this study indicate synergistic interactions among contractile factors affect ovulatory outcome, while the role of EDNRB alone in the process of ovulation requires further investigation.

Soluble Prorenin Receptor Increases Blood Pressure in High Fat-Fed Male Mice.

Obesity-related hypertension is a major public health concern. We recently demonstrated that plasma levels of the soluble form of the prorenin receptor (sPRR) were elevated in obesity-associated hypertension. Therefore, in the present study, we investigated the contribution of sPRR to blood pressure (BP) elevation in the context of obesity. High fat-fed C57BL/6 male mice were infused with vehicle or sPRR (30 µg/kg per day) via subcutaneously implanted osmotic minipump for 4 weeks. BP parameters were recorded using radiotelemetry devices. Male mice infused with sPRR exhibited higher systolic BP and mean arterial pressure and lower spontaneous baroreflex sensitivity than mice infused with vehicle. To define mechanisms involved in systolic BP elevation, mice were injected with an AT1R (Ang II [angiotensin II] type 1 receptor) antagonist (losartan), a muscarinic receptor antagonist (atropine), a ß-adrenergic antagonist (propranolol), and a ganglionic blocker (chlorisondamine). Losartan did not blunt sPRR-induced elevation in systolic BP. Chlorisondamine treatment exacerbated the decrease in mean arterial pressure in male mice infused with sPRR. These results demonstrated that sPRR induced autonomic nervous dysfunction. Interestingly, plasma leptin levels were increased in high fat-fed C57BL/6 male mice infused with sPRR. Overall, our results indicated that sPRR increased systolic BP through an impairment of the baroreflex sensitivity and an increase in the sympathetic tone potentially mediated by leptin in high fat-fed C57BL/6 male mice.